Chemistry Reference
In-Depth Information
5.5.3 PDF
Compound
6
is one of numerous fragments identified in an in-house NMR screen of PDF.
The solution structure of PDF in complex with
6
was determined by restrained simulated
annealing
[
27
]
using data derived from standard three-dimensional NMR techniques. The
average structure of the resultant ensemble of NMR structures was calculated and subjected
to unrestrained energy minimization; this structure served as the 'target pose' for the NOE
matching simulations. To generate the required input files for NOEmatching, a list of PDF/
6
NOEs was derived from distances calculated using the PDF/
6
complex using a distance
cutoff of 4.5 Å (a conservative upper bound estimate compared to our real experimental
NOESYdata) and using the BMRB average chemical shifts for the simulated 'experimental'
chemical shifts. The simulated NOE list for this complex contained a total of 62 peaks,
which were clustered into 48 protein
1
H
13
C groups. Trial binding poses were generated
with
Poser
. The compound binding site was defined as the active site of the protein; the
'posing box' was expanded by 1 Å in all coordinate axes. For each of the three ligand
conformers (generated with Omega), 485 968 896 poses were generated and evaluated by
Poser
, with 50 792 poses being retained. From this set of poses, 1000 poses were selected
by random sampling for scoring by NOE matching. The RMSDs of the retained trial poses
to the target pose ranged from 0.96 to 5.75 Å. The NOE matching protocol was run using
BMRB-predicted chemical shifts.
The results obtained from applying NOE matching to PDF/
6
are shown in Figure 5.8.
The pose with the minimum COST value has an RMSD of 1.02 Å to the target pose. The
pose with the closest RMSD to the target pose itself ranks 6 out of 1000 poses.
(a)
(b)
3000
2500
2000
1500
1000
500
0
0
1
2
3
RMSD to Target
4
5
6
Figure 5.8
(A) COST versus the RMSD (Å) to the target pose for PDF/
6
. The predicted pro-
tein chemical shifts were set to the corresponding BMRB average values. The 3D X-filtered
NOESY spectrum used as input for NOE matching was simulated from the target structure.
(B) Superposition of target pose and the minimum cost pose (dark gray) from (A).
Two predominant binding modes were scored as having a low COST by NOE matching.
These binding modes can be observed in Figure 5.8A as the lowest COST poses (around
1 Å from the target pose) and a second binding mode whose members are
∼
3.4 Å from the
∼
target pose. These two binding modes represent poses that are
180º flipped with respect
